title: Method qualification and application of diffusion interaction parameter and virial coefficient.
abstract: This research focused on evaluation and application of two methods in studying weak protein-protein interactions, i.e. diffusion interaction parameter (KD) and second virial coefficient (B22), both of which are first-order coefficients of protein interactions. Although the plate-based KD method successfully distinguished KD values with relatively large difference in a pH ranging study, it failed to make a consistent statistical decision to determine close interactions as shown by the comprehensive ANOVA analysis. We also validated the DLS-based B22 method by using a model protein lysozyme. The dramatic change of solution appearance for lysozyme as a function of NaCl concentration highlighted the importance of B22 in understanding protein interactions. Moreover, B22 measurement for a MAb fragment suggested a more repulsive protein interaction in histidine buffer than in citrate buffer. The coefficient of variation was <10% when B22 was on an order of magnitude of 10(-4) L mmol/g(2) in contrast to >30% when it approached 10(-5) L mmol/g(2). In this research, we also made an attempt to study protein-protein interactions in concentrated MAb fragment solutions (e.g. >50 mg/mL). Our data suggested that such interactions could be empirically modeled by high-order virial expansions.
authors.affiliation: Sterile Product Development Group, Bioprocess Development, Merck Sharp & Dohme Corp., 181 Passaic Avenue, Summit, NJ 07901, United States. Electronic address: shuai.shi@merck.com.
authors.forename: Shuai
authors.initials: S
authors.lastname: Shi
authors.forename: Makiko
authors.initials: M
authors.lastname: Uchida
authors.forename: Jason
authors.initials: J
authors.lastname: Cheung
authors.forename: Valentyn
authors.initials: V
authors.lastname: Antochshuk
authors.forename: Mohammed
authors.initials: M
authors.lastname: Shameem
mesh_terms: D000818:Animals; D004058:Diffusion; D004527:Egg Proteins / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D009113:Muramidase / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D015203:Reproducibility of Results
chemical_list: D004527:Egg Proteins; D011506:Proteins; D009113:Muramidase
keywords: ANOVA; Diffusion interaction parameter; Protein–protein interaction; Variability; Virial coefficient